Specimen Collection

INDICATIONS FOR REQUESTING TEST

1. Chronic destructive disease of the upper airways
2. Pulmonary nodules, non-malignant
3. Subglottic stenosis of the trachea
4. Pulmonary-renal syndrome
5. Glomerulonephritis
6. Suspected vasculitis
7. Mononeuritis multiplex
8. Retro-orbital mass

NOTE: This testing can be requested as routine where there is a low pre-test probability of a small vessel vasculitis (SVV) or as an urgent request where there is a high probability of a small vessel vasculitis and delivery of results will immediately change therapy for the patient.

There are differences in registration between "routine" and "urgent" requesting as outlined below:-

3.5 mL SST Serum (Preferred)

4 mL Plain Blood

250 uL Microsample Serum

(or 2 x 1mL plain micro-collects)

Reference Intervals

PR3-ANCA and MPO-ANCA

Due to (a) the use of arbitrary reporting units and (b) lack of harmonisation across diagnostic assays for MPO and PR3 antibodies it is strongly recommended that comparison of reported absolute numeric values across different assays / platforms be avoided for patient monitoring purposes.

Normal : < 20 AU/mL
Borderline: 20-25 AU/mL
Abnormal: >25 AU/mL

Uncertainty of Measurement: 20% (both).

Turnaround Time:

Turnaround time (f rom Specimen arrival time to Reporting) :

• For Routine requests : 2-3 days,
• For Urgent requests : 3.5 hours (please refer to the "Assay method" Section for the details).
  

Assay Method

Routine requests

Serum are screened by indirect immunofluorescence (IIF), and specific antibody tests for PR-3 and MPO are performed on positive samples using luminex methodology.

Labplus Order Code: Alpha Code: NEUT, Numeric Code:7155

If there is nothing documented on the request form to indicate either "rapid" or "urgent" then treat as a routine request.

Turnaround time: 2-3 days (Negative result), within 1 week for IIF reactive sera IIF C-ANCA IIF patterns are strongly associated with GPA [Granulomatosis with Polyangiitis, previously Wegener?s Granulomatosis] (positive in 85 to 100% of patients with active generalised disease, less frequently in patients with limited disease). The target antigen giving a C-ANCA IIF pattern is PR3 (Proteinase 3).

P-ANCA IIF patterns are seen in the following SVV's:- MPA microscopic polyangiitis, idiopathic necrotising and crescentic glomerulonephritis (pauci-immune GN), EGPA (Eosinophilic Granulomatosis with Polyangiitis, previously Churg-Strauss syndrome). However, they can also be present in several non-SVV related diseases. The target antigen giving a P-ANCA pattern in a SVV disease is MPO (Myeloperoxidase).

In patients not under treatment with a known SVV, a negative IIF result (both patterns) excludes an ANCA-mediated vasculitic disease.

Urgent requests

This assay is restricted and must be approved prior to the assay being performed. The approval process is:
Urgent vasculitis requests during normal working hours must be pre-approved by either, the Immunopathologist, Immunopathology registrar or a senior member of the testing team in that order of preference. Any request not pre-approved will be treated as routine.

To ensure prioritised processing notify Section Leader (VIM - Serology) or their delegated authority on xtn 23006 22104 / 22103.

Urgent vasculitis requests outside normal working hours will only be performed once the Consultant of the requesting team has contacted the Immunopathologist and reached agreement that the test is required out of hours. Requests not from the requesting team's Consultant and/or not approved by the Immunopathologist will not be actioned .

Results will be rung through by the testing scientist. Any discussions on the relationship of the results to clinical diagnosis will need to be directed to the Immunopathologist or their registrar.

No IIF testing will be performed. ANCA-MPO and ANCA-PR3 will be tested.

Please note that GBM testing is not part of the urgent ANCA testing and needs to be specifically requested (refer GBM as the search term in the on-line Test Guide)

Out of hours includes:
- Weekends
- 17.00-08.00 on weekdays
- Public holidays

Diagnostic Use and Interpretation

Anti-neutrophilic cytoplasmic antibodies (ANCA) represent a group of autoantibodies directed towards the cytoplasmic components of the neutrophilic granulocytes and monocytes. Using IFA techniques, two main patterns are recognised (a) a cytoplasmic [C-ANCA] and (b) perinuclear [P-ANCA] type. The main antigen for C-ANCA is proteinase 3 [PR3], a serine proteinase present in primary granules. P-ANCA patterns in general are due to antibodies directed against myeloperoxidase [MPO]. Problems exist in the interpretation of IFA patterns, due to antibodies versus lactoferrin, elastase, cathepsin-G and lysozyme resulting in P-ANCA patterns. Granulocyte specific antinuclear antibodies (GS-ANA) will also result in IFA patterns indistinguishable from P-ANCA. Use of MPO-ANCA and PR3-ANCA in conjunction with the classical ANCA IFA and ANA IFA techniques allows a greater level of discrimination when considering the clinical implication of ANCA results.

Documented specific clinical indications for ANCA is presented in the following Table. MPO-ANCA and PR3-ANCA are reliable serologic markers in the diagnosis of vasculitidies. PR3-ANCA is the classical autoantigen in GPA with a clinical specificity of greater than 95%. MPO is mainly present in MPA (70%).

Disease	ANCA IFA pattern	Target Antigen
Systemic Vasculitic
GPA	C-ANCA, rare P-ANCA	PR3, rare MPO
MPA	C-ANCA, P-ANCA	PR3, MPO
EGPA	P-ANCA	MPO
Polyarteritis nodosa	Rare ANCA	Rare MPO and PR3
Unclassified vasculitidies	Rare	No PR3 and MPO
Collagen / Rheumatic
RA	GS-ANA, P-ANCA	Unknown, ANA, rare MPO, lactoferrin
SLE	P-ANCA	Rare MPO, lactoferrin
Others
Ulcerative colitis	P-ANCA	Cathepsin G, lactoferrin and other unknown antigens
Morbus Crohn
Chronic Hepatitis

References:

1. FIDIS - Vasculitis product insert.

Contact Information

For further information contact the laboratory, (09) 307 4949 ext 22103 or:
Associate Professor Rohan Ameratunga , Immunopathologist: Locator 93-5724,

Dr Richard Steele , or The LabPLUS Immunology Team

Clinical

Associate Professor Rohan Ameratunga , Immunopathologist: Locator 93-5724,
Dr Richard Steele , or The LabPLUS Immunology Team

Technical

Paul Austin, Section Lead 022 632 1554
Helena Thompson-Faiva, Serology 2IC, Technical Specialist x 22104
Rong Zhou, Technical Specialist x 22103
Yulia Hwang, Technical Specialist x 22103

Last updated at 12:39:20 28/06/2023